It was previously reported that several retrovirus and lentivirus particles contained a novel topoisomerase I (topo I) activity and that the specific inhibitor of this enzyme, camptothecin, could block HIV infection of H9 cells at concentrations which were not cytotoxic. In an effort to identify derivatives of camptothecin which might exhibit an improved antiviral activity with lower cytotoxicity, a panel of 20 derivatives of CPT were screened for their ability to inhibit HIV infection in tissue culture. Results indicated that derivatives previously known to be ineffective inhibitors of topoisomerase I activity showed no inhibition of HIV. Derivatives capable of inhibiting topo I activity in vitro were able to inhibit HIV infection of H9, but all showed significant cytotoxicity. None of the derivatives tested were significantly more effective antiviral agents than the parent compound camptothecin.